University of Nebraska - Lincoln DigitalCommons@University of Nebraska - Lincoln Papers in Plant Pathology Plant Pathology Department 2008 Reclassification of Subspecies of Acidovorax avenae as A. Avenae (Manns 1905) emend., A. cattleyae (Pavarino, 1911)comb.nov., A. citrulli Schaad et al.,1978)comb.nov., and proposal of A. oryzae sp. nov. Norman Schaad USDA-ARS Elena Postnikova USDA-ARS Aaron Sechler USDA-ARS Larry E. Claflin Kansas State University Anne K. Vidaver University of Nebraska-Lincoln, [email protected] SeeFollow next this page and for additional additional works authors at: https:/ /digitalcommons.unl.edu/plantpathpapers Part of the Plant Pathology Commons Schaad, Norman; Postnikova, Elena; Sechler, Aaron; Claflin, Larry E.; Vidaver, Anne K.; Jones, Jeffrey B.; Agarkova, Irina V.; Ignatov, Alexander; Dickstein, Ellen; and Ramundo, Bruce A., "Reclassification of Subspecies of Acidovorax avenae as A. Avenae (Manns 1905) emend., A. cattleyae (Pavarino, 1911)comb.nov., A. citrulli Schaad et al.,1978)comb.nov., and proposal of A. oryzae sp. nov." (2008). Papers in Plant Pathology. 133. https://digitalcommons.unl.edu/plantpathpapers/133 This Article is brought to you for free and open access by the Plant Pathology Department at DigitalCommons@University of Nebraska - Lincoln. It has been accepted for inclusion in Papers in Plant Pathology by an authorized administrator of DigitalCommons@University of Nebraska - Lincoln. Authors Norman Schaad, Elena Postnikova, Aaron Sechler, Larry E. Claflin, Anne K. Vidaver, Jeffrey B. Jones, Irina V. Agarkova, Alexander Ignatov, Ellen Dickstein, and Bruce A. Ramundo This article is available at DigitalCommons@University of Nebraska - Lincoln: https://digitalcommons.unl.edu/ plantpathpapers/133 ARTICLE IN PRESS Systematic and Applied Microbiology 31 (2008) 434–446 www.elsevier.de/syapm Reclassification of subspecies of Acidovorax avenae as A. Avenae (Manns 1905) emend., A. cattleyae (Pavarino, 1911) comb. nov., A. citrulli Schaad et al., 1978) comb. nov., and proposal of A. oryzae sp. nov. Norman W. Schaada,Ã,1, Elena Postnikovaa,1, Aaron Sechlera, Larry E. Claflinb, Anne K. Vidaverc, Jeffrey B. Jonesd, Irina Agarkovac, Alexander Ignatove, Ellen Dicksteind, Bruce A. Ramundob aARS-USDA, Foreign Disease-Weed Science Research Unit, 1301 Ditto Avenue, Ft. Detrick, MD 21702, USA bDepartment of Plant Pathology, Kansas State University, Manhattan, KS, USA cDepartment of Plant Pathology, University of Nebraska, Lincoln, NB, USA dDepartment of Plant Pathology, University of Florida, USA eCenter ‘‘Bioengineering’’ RAS, Moscow, Russia Abstract The bacterium Acidovorax avenae causes disease in a wide range of economically important monocotyledonous and dicotyledonous plants, including corn, rice, watermelon, anthurium, and orchids. Genotypic and phenotypic relatedness among strains of phytopathogenic A. avenae subsp. avenae, A. avenae subsp. citrulli, A. avenae subsp. cattleyae and A. konjaci, as well as all other Acidovorax species, including A. facilis, the type strain of Acidovorax, was determined. The 16s rDNA sequencing confirmed previous studies showing the environmental species to be very distant from the phytopathogenic species. DNA/DNA reassociation assays on the different strains of A. avenae revealed four (A, B, C, and D) distinct genotypes. Taxon A included six A. avenae subsp. avenae strains from corn that had a mean reciprocal similarity of 81%; taxon B included six A. avenae subsp. avenae strains from rice that had a mean reciprocal similarity of 97%; taxon C contained 11 A. avenae subsp. citrulli strains from cucurbits (cantaloupe, watermelon, and pumpkin) that had a mean reciprocal similarity of 88%, and taxon D contained four A. avenae subsp. cattleyae strains from orchids that had a mean similarity of 98%. The mean reciprocal relatedness between taxa A, B, C, and D was less than 70%. Sequence analysis of 16S rDNA and the 16S–23S rDNA internally transcribed spacer region, as well as AFLP analysis, revealed the same four taxa. All four were easily differentiated phenotypically from each other and from all other recognized Acidovorax species. Strains of A. avenae did not contain 3-hydroxyoctanoic acid, which was found in all other species. On the basis of these and previous genetic and phenotypic results, we propose an emendation of the species A. avenae. A. avenae subsp. citrulli (C strains) and A. avenae subsp.cattleyae(D strains) should be elevated to species rank as A. citrulli and A. cattleyae, respectively. We further propose a new taxon for the B strains, A. oryzae sp. nov. with FC-143T ¼ ICPB 30003T ¼ ICMP 3960T ¼ ATCC 19882T as the type strain. Published by Elsevier GmbH. Keywords: Acidovorax; Taxonomy; 16S and ITS sequencing; AFLP; Phenotypic tests; DNA/DNA reassociation; Emendation ÃCorresponding author. E-mail address: [email protected] (N.W. Schaad). 1Authors contributed equally to this work. 0723-2020/$ - see front matter Published by Elsevier GmbH. doi:10.1016/j.syapm.2008.09.003 ARTICLE IN PRESS N.W. Schaad et al. / Systematic and Applied Microbiology 31 (2008) 434–446 435 Introduction to be part of the ‘‘acidovorans’’ DNA–rRNA homology group [7]. Acidovorax avenae subsp. avenae (Pseudomonas ave- Based on DNA/DNA and DNA–rRNA homologies nae) and A. avenae subsp. citrulli (synonymous with [5] and phenotypic assays, Willems et al. [55] proposed P. pseudoalcaligenes subsp. citrulli) have emerged world- that the above ‘‘generically misnamed phytopatho- wide as serious pathogens on corn (Zea mays L.) [40] genic pseudomonads’’ be transferred to the newly and watermelon (Citrullus lanatus (Thunb.) Matsumura proposed genus Acidovorax [54]. Strains of P. avenae and Nakai) [42], respectively. Pseudomonas avenae, [P. rubrilineans] from corn, sugar cane, Indian shot originally described in 1909 as the causal agent of leaf (Canna indica), teosinte, finger millet, and ‘‘P. setariae’’ blight of oats (Avenae sativa L.) [31], causes disease from rice were all classified as A. avenae subsp. avenae. under conditions of high rainfall and high temperatures Strains of P. pseudoalcaligenes subsp. citrulli from in numerous species of the family Graminaceae, includ- Cucurbitaceae hosts were classified as A. avenae subsp. ing wheat (Triticum aestivum L.), finger millet (Eleusine citrulli; strains of P. cattleyae from orchids were coracana (L.) Gaertn.), Italian millet (Setaria italica (L.) renamed as A. avenae subsp. cattleyae; and strains of Beauv.), pearl millet (Pennisetum glaucum (L.) R. Br.), P. pseudoalcaligenes subsp. konjaci [15] from konjac and proso millet (Panicum miliaceum L.) [4,9,32,33]. were classified as A. konjaci [55]. Recently two new Additional hosts include tea (Thea sinensis L.), barley phytopathogenic species of Acidovorax, A. anthurii on (Hordeum vulgare L.), mountain brome (Bromus car- anthurium (Anthurium palmatum (L.) G. Don) [11] and inatus Hook. & Arn.), rescue grass (B. catharticus Vahl), A. valerianellae on lambs’ lettuce (Valerianella locusta vasey grass (Paspalum urvillei Steud.), teosinte (Zea (L.) Lat.) [12], as well as a denitrifying species, A. caeni mexicana (Schrad.) Kuntze), and sugarcane (Saccharum from activated sludge [18], have been described. officinarum L.) [2,13,17,49,50]. P. alboprecipitans,a The purpose of this study was to re-evaluate the pathogen of foxtail (Chaetochloa lutescens (Weigel) phylogenetic relatedness among the above phytopatho- Stuntz) [36], was later shown to be synonymous with genic A. avenae strains and between A. avenae and the the oats and corn pathogen, P. avenae [40]. Addition- phytopathogenic A. konjaci, A. anthurii,andA. valer- ally, the red stripe disease of sugarcane caused by ianellae. A polyphasic taxonomic study revealed con- P. rubrilineans [17] was shown to be synonymous with siderable differences among the different subspecies of P. avenae [35]. More recently A. avenae subsp. avenae A. avenae and between the other species of Acidovorax. has emerged as a serious pathogen of rice (Oryzae sativa Therefore, we propose a new species, A. oryzae for the L.) [16,24,26,27,34]. The causal agent of bacterial stripe rice strains of A. avenae. We also propose A. avenae of rice also has been referred to as ‘‘[P.] setariae’’ [14] subsp. citrulli and A. avenae subsp. cattleyae be elevated and P. panici [34], however, neither epithet has been to species rank as A. citrulli comb. nov. and A. cattleyae validly published [56]. A pathogen of orchids (Cattleya, comb. nov. Dendrobium, Oncidium, and Phalaenopsis), P. cattleyae, was described in 1946 [1]. A pathogen of konjac (Amorphophallus konjac K. Koch.), A. konjaci, was Materials and methods described in 1983 [15]. All of these phytopathogenic bacteria were described previously as a heterogenous group of non-fluorescent, oxidase positive pseudomo- Source of strains and confirmation of identity nads, which accumulated poly-b-hydroxybutyrate All the strains used in this study were obtained from (PHB) and utilized D-arabinose [38]. the International Collection of Phytopathogenic Bacteria Schaad et al. [41] described a non-fluorescent pseu- (ICPB) maintained at the USDA, ARS Foreign Disease- domonad as the causal agent of a seedling blight of Weed Science Research Unit (FDWSRU), Fort Detrick, watermelon. The bacterium differed phenotypically from other known plant pathogenic pseudomonads MD or from other recognized culture collections (Table S-1). Each culture was streaked onto yeast and was classified as P. pseudoalcaligenes subsp. citrulli extract-dextrose CaCO (YDC) agar [23] and